Proteomic Profiling of Burkholderia thailandensis During Host Infection Using Bio-Orthogonal Noncanonical Amino Acid Tagging (BONCAT)
Burkholderia pseudomallei and B. mallei are the causative agents of melioidosis and glanders, respectively, and are often fatal to humans and animals. Owing to the high fatality rate, potential for spread by aerosolization, and the lack of efficacious therapeutics, B. pseudomallei and B. mallei are...
| Main Authors: | , , , , , , |
|---|---|
| Format: | Article |
| Language: | English |
| Published: |
Frontiers Media S.A.
2018-10-01
|
| Series: | Frontiers in Cellular and Infection Microbiology |
| Subjects: | |
| Online Access: | https://www.frontiersin.org/article/10.3389/fcimb.2018.00370/full |
| _version_ | 1818137565995204608 |
|---|---|
| author | Magdalena Franco Patrik M. D'haeseleer Steven S. Branda Megan J. Liou Yasmeen Haider Brent W. Segelke Sahar H. El-Etr |
| author_facet | Magdalena Franco Patrik M. D'haeseleer Steven S. Branda Megan J. Liou Yasmeen Haider Brent W. Segelke Sahar H. El-Etr |
| author_sort | Magdalena Franco |
| collection | DOAJ |
| description | Burkholderia pseudomallei and B. mallei are the causative agents of melioidosis and glanders, respectively, and are often fatal to humans and animals. Owing to the high fatality rate, potential for spread by aerosolization, and the lack of efficacious therapeutics, B. pseudomallei and B. mallei are considered biothreat agents of concern. In this study, we investigate the proteome of Burkholderia thailandensis, a closely related surrogate for the two more virulent Burkholderia species, during infection of host cells, and compare to that of B. thailandensis in culture. Studying the proteome of Burkholderia spp. during infection is expected to reveal molecular mechanisms of intracellular survival and host immune evasion; but proteomic profiling of Burkholderia during host infection is challenging. Proteomic analyses of host-associated bacteria are typically hindered by the overwhelming host protein content recovered from infected cultures. To address this problem, we have applied bio-orthogonal noncanonical amino acid tagging (BONCAT) to B. thailandensis, enabling the enrichment of newly expressed bacterial proteins from virtually any growth condition, including host cell infection. In this study, we show that B. thailandensis proteins were selectively labeled and efficiently enriched from infected host cells using BONCAT. We also demonstrate that this method can be used to label bacteria in situ by fluorescent tagging. Finally, we present a global proteomic profile of B. thailandensis as it infects host cells and a list of proteins that are differentially regulated in infection conditions as compared to bacterial monoculture. Among the identified proteins are quorum sensing regulated genes as well as homologs to previously identified virulence factors. This method provides a powerful tool to study the molecular processes during Burkholderia infection, a much-needed addition to the Burkholderia molecular toolbox. |
| first_indexed | 2024-12-11T09:58:20Z |
| format | Article |
| id | doaj.art-afc40db9ae9f4e47985a500a2126d331 |
| institution | Directory Open Access Journal |
| issn | 2235-2988 |
| language | English |
| last_indexed | 2024-12-11T09:58:20Z |
| publishDate | 2018-10-01 |
| publisher | Frontiers Media S.A. |
| record_format | Article |
| series | Frontiers in Cellular and Infection Microbiology |
| spelling | doaj.art-afc40db9ae9f4e47985a500a2126d3312022-12-22T01:12:11ZengFrontiers Media S.A.Frontiers in Cellular and Infection Microbiology2235-29882018-10-01810.3389/fcimb.2018.00370409343Proteomic Profiling of Burkholderia thailandensis During Host Infection Using Bio-Orthogonal Noncanonical Amino Acid Tagging (BONCAT)Magdalena Franco0Patrik M. D'haeseleer1Steven S. Branda2Megan J. Liou3Yasmeen Haider4Brent W. Segelke5Sahar H. El-Etr6Lawrence Livermore National Laboratory, Livermore, CA, United StatesLawrence Livermore National Laboratory, Livermore, CA, United StatesSandia National Laboratories, Livermore, CA, United StatesLawrence Livermore National Laboratory, Livermore, CA, United StatesLawrence Livermore National Laboratory, Livermore, CA, United StatesLawrence Livermore National Laboratory, Livermore, CA, United StatesLawrence Livermore National Laboratory, Livermore, CA, United StatesBurkholderia pseudomallei and B. mallei are the causative agents of melioidosis and glanders, respectively, and are often fatal to humans and animals. Owing to the high fatality rate, potential for spread by aerosolization, and the lack of efficacious therapeutics, B. pseudomallei and B. mallei are considered biothreat agents of concern. In this study, we investigate the proteome of Burkholderia thailandensis, a closely related surrogate for the two more virulent Burkholderia species, during infection of host cells, and compare to that of B. thailandensis in culture. Studying the proteome of Burkholderia spp. during infection is expected to reveal molecular mechanisms of intracellular survival and host immune evasion; but proteomic profiling of Burkholderia during host infection is challenging. Proteomic analyses of host-associated bacteria are typically hindered by the overwhelming host protein content recovered from infected cultures. To address this problem, we have applied bio-orthogonal noncanonical amino acid tagging (BONCAT) to B. thailandensis, enabling the enrichment of newly expressed bacterial proteins from virtually any growth condition, including host cell infection. In this study, we show that B. thailandensis proteins were selectively labeled and efficiently enriched from infected host cells using BONCAT. We also demonstrate that this method can be used to label bacteria in situ by fluorescent tagging. Finally, we present a global proteomic profile of B. thailandensis as it infects host cells and a list of proteins that are differentially regulated in infection conditions as compared to bacterial monoculture. Among the identified proteins are quorum sensing regulated genes as well as homologs to previously identified virulence factors. This method provides a powerful tool to study the molecular processes during Burkholderia infection, a much-needed addition to the Burkholderia molecular toolbox.https://www.frontiersin.org/article/10.3389/fcimb.2018.00370/fullBurkholderiaBONCATorthogonal amino acid labelingintracellular pathogenhost infectionprotein enrichment |
| spellingShingle | Magdalena Franco Patrik M. D'haeseleer Steven S. Branda Megan J. Liou Yasmeen Haider Brent W. Segelke Sahar H. El-Etr Proteomic Profiling of Burkholderia thailandensis During Host Infection Using Bio-Orthogonal Noncanonical Amino Acid Tagging (BONCAT) Frontiers in Cellular and Infection Microbiology Burkholderia BONCAT orthogonal amino acid labeling intracellular pathogen host infection protein enrichment |
| title | Proteomic Profiling of Burkholderia thailandensis During Host Infection Using Bio-Orthogonal Noncanonical Amino Acid Tagging (BONCAT) |
| title_full | Proteomic Profiling of Burkholderia thailandensis During Host Infection Using Bio-Orthogonal Noncanonical Amino Acid Tagging (BONCAT) |
| title_fullStr | Proteomic Profiling of Burkholderia thailandensis During Host Infection Using Bio-Orthogonal Noncanonical Amino Acid Tagging (BONCAT) |
| title_full_unstemmed | Proteomic Profiling of Burkholderia thailandensis During Host Infection Using Bio-Orthogonal Noncanonical Amino Acid Tagging (BONCAT) |
| title_short | Proteomic Profiling of Burkholderia thailandensis During Host Infection Using Bio-Orthogonal Noncanonical Amino Acid Tagging (BONCAT) |
| title_sort | proteomic profiling of burkholderia thailandensis during host infection using bio orthogonal noncanonical amino acid tagging boncat |
| topic | Burkholderia BONCAT orthogonal amino acid labeling intracellular pathogen host infection protein enrichment |
| url | https://www.frontiersin.org/article/10.3389/fcimb.2018.00370/full |
| work_keys_str_mv | AT magdalenafranco proteomicprofilingofburkholderiathailandensisduringhostinfectionusingbioorthogonalnoncanonicalaminoacidtaggingboncat AT patrikmdhaeseleer proteomicprofilingofburkholderiathailandensisduringhostinfectionusingbioorthogonalnoncanonicalaminoacidtaggingboncat AT stevensbranda proteomicprofilingofburkholderiathailandensisduringhostinfectionusingbioorthogonalnoncanonicalaminoacidtaggingboncat AT meganjliou proteomicprofilingofburkholderiathailandensisduringhostinfectionusingbioorthogonalnoncanonicalaminoacidtaggingboncat AT yasmeenhaider proteomicprofilingofburkholderiathailandensisduringhostinfectionusingbioorthogonalnoncanonicalaminoacidtaggingboncat AT brentwsegelke proteomicprofilingofburkholderiathailandensisduringhostinfectionusingbioorthogonalnoncanonicalaminoacidtaggingboncat AT saharheletr proteomicprofilingofburkholderiathailandensisduringhostinfectionusingbioorthogonalnoncanonicalaminoacidtaggingboncat |